title: MMP20 and KLK4 activation and inactivation interactions in vitro.
abstract: OBJECTIVE
  Enamelysin (MMP20) and kallikrein 4 (KLK4) are believed to be necessary to clear proteins from the enamel matrix of developing teeth. MMP20 is expressed by secretory stage ameloblasts, while KLK4 is expressed from the transition stage throughout the maturation stage. The aim of this study is to investigate the activation of KLK4 by MMP20 and the inactivation of MMP20 by KLK4.
  
  
  METHODS
  Native pig MMP20 (pMMP20) and KLK4 (pKLK4) were isolated directly from enamel scrapings from developing molars. Recombinant human proKLK4 (rh-proKLK4) was activated by incubation with pMMP20 or recombinant human MMP20 (rhMMP20), and the resulting KLK4 activity was detected by zymography. Reaction products were isolated by reverse-phase high performance liquid chromatography (RP-HPLC), and their N-termini characterized by Edman degradation. The pMMP20 was incubated with pKLK4 under mildly acidic or under physiologic conditions, and enzyme activity was analyzed by zymography. The catalytic domain of rhMMP20 was incubated with pKLK4 or recombinant human KLK4 (rhKLK4) and the digestion products were characterized by zymography and Edman degradation.
  
  
  RESULTS
  Both pMMP20 and rhMMP20 activated rh-proKLK4 by cleaving at the propeptide-enzyme junction used in vivo. The pMMP20 was inactivated by pKLK4 under physiologic conditions, but not under mildly acidic conditions. Both pKLK4 and rhKLK4 cleaved MMP20 principally at two sites in the catalytic domain of MMP20.
  
  
  CONCLUSIONS
  MMP20 activates proKLK4 and KLK4 inactivates MMP20 in vitro, and these actions are likely to occur during enamel formation in vivo.
authors.affiliation: Department of Biochemistry and Molecular Biology, School of Dental Medicine, Tsurumi University, 2-1-3 Tsurumi, Tsurumi-ku, Yokohama 230-8501, Japan. Electronic address: yamakoshi-y@tsurumi-u.ac.jp.
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Yamakoshi
authors.forename: James P
authors.initials: JP
authors.lastname: Simmer
authors.forename: John D
authors.initials: JD
authors.lastname: Bartlett
authors.forename: Takeo
authors.initials: T
authors.lastname: Karakida
authors.forename: Shinichiro
authors.initials: S
authors.lastname: Oida
mesh_terms: D000565:Ameloblasts / Q000201:enzymology*; D000566:Amelogenesis / Q000502:physiology*; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D003746:Dental Enamel Proteins / Q000378:metabolism*; D004492:Edetic Acid; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D006851:Hydrochloric Acid; D007610:Kallikreins / Q000302:isolation & purification / Q000378:metabolism*; D053519:Matrix Metalloproteinase 20 / Q000302:isolation & purification / Q000378:metabolism*; D061965:Matrix Metalloproteinase Inhibitors / Q000378:metabolism*; D008963:Molar / Q000201:enzymology; D009943:Organophosphorus Compounds; D011994:Recombinant Proteins / Q000378:metabolism; D034421:Sus scrofa
chemical_list: C043517:2-(4-isothiocyanatophenoxy)-1,3,2-dioxaphosphinene 2-oxide; D003746:Dental Enamel Proteins; D061965:Matrix Metalloproteinase Inhibitors; D009943:Organophosphorus Compounds; D011994:Recombinant Proteins; D004492:Edetic Acid; D007610:Kallikreins; C120801:kallikrein 4; D053519:Matrix Metalloproteinase 20; D006851:Hydrochloric Acid
keywords: Enamel; KLK4; MMP20; Proteases; Tooth